Total poly A containing mRNA isolated from Angler fish and cold islets of Langerhans will be translated in the wheat germ cell-free protein synthesizing system to characterize putative preproglucagon and preproinsulin and to utilize this system for testing predictions of the "Signal Hypothesis." The precursors will be identified by antibody precipitation, SDS-gel electrophoresis and peptide mapping. The complete amino acid sequence of the N-terminal signal regions of the molecules, labelled with 4 or 5 radioactive amino acids, will be determined by automatic Edman degradation. Mammalian stripped microsomal membranes will be added to the cell-free system co-translationally to determine and identify the processes and segregated products of proglucagon. The fate and ultimate localization of the signal peptide following its removal from preproinsulin and preproglucagon will be investigated by isolating the exogenous microsomal vesicles from the translation system and characterizing the segregated products by gel electrophoresis, gel filtration and automatic Edman degradation. If the signal is not immediately degraded its subsequent localization will be determined using cell fractionation and experiments will be performed to determine if the signal is secreted. The post-translational cleavage reactions in the information of glucagon from preproglucagon will be investigated by adding Golgi membranes and crude secretory granules to the cell-free system containing stripped microsomal vesicles. The role of these organelles in converting proglucagon to glucagon will be examined by identifying the intermediate products generated and the localization of the enzymatic activities responsible for processing proglucagon will be investigated.